Current modes of therapy for the prevention and/or treatment of apoptosis-related and neurodegenerative diseases, ischemic conditions, COPD, macular degeneration, microvascular diseases and ototoxic conditions are unsatisfactory and there is a need therefore to develop novel compounds for this purpose. The present invention is focused on processes for identifying such compounds. All the diseases and indications disclosed herein, as well as other diseases and conditions disclosed in coassigned PCT Application No. WO06/023544A2 may also be treated by the novel compounds of this invention.
RTP801
Gene RTP801, was first disclosed in commonly assigned U.S. Pat. Nos. 6,455,674, 6,555,667, and 6,740,738, including the RTP801 polynucleotide and polypeptide, and antibodies directed toward the polypeptide. RTP801 represents a unique gene target for hypoxia-inducible factor-1 (HIF-1) that may regulate hypoxia-induced pathogenesis independent of growth factors such as VEGF.
The Following Patent Applications and Publications Provide Background Information:
PCT Publication No. WO 2001070979 relates to nucleic acid markers, which are overexpressed in ovarian cancer cells.
U.S. Pat. No. 6,673,549 discloses a combination comprising cDNAs that are differentially expressed in response to steroid treatment.
U.S. Patent Publication No. 2003165864 relates to cDNAs that are differentially expressed in cells treated with a DNA demethylating agent.
U.S. Patent Publication No. 2003108871 relates to a composition comprising several cDNAs that are differentially expressed in treated human C3A liver cell cultures.
U.S. Patent Publication No. 2002119463 discloses a new composition, useful for treating and diagnosing prostate cancer, said composition comprising human cDNAs that are differentially expressed in prostate cancer.
PCT Publication No. WO 2004018999 discloses a method for assessing, characterizing, monitoring, preventing and treating cervical cancer.
European Patent No. EP 1394274 relates to a method of testing for bronchial asthma or chronic obstructive pulmonary disease by comparing the expression level of a marker gene in a biological sample from a subject with the expression level of the gene in a sample from a healthy subject.
PCT Publication No. WO 2002101075 relates to an isolated nucleic acid molecule useful for detecting, characterizing, preventing and treating human cervical cancers.
PCT Publication No. WO 2003010205 relates to inhibiting angiogenesis for treating wound healing, retinopathy, ischemia, inflammation, microvasculopathy, bone healing and skin inflammation.
PCT Publication No. WO 2002046465 relates to identifying a gene involved in disease for treating hypoxia- regulated conditions.
PCT Publication No. WO 2002031111 relates to polypeptides and their encoded proteins, and many uses therefore are provided.
PCT Publication No. WO 2001012659 relates to nucleic acids useful in recombinant DNA methodologies.
PCT Publication No. WO 2001077289 discloses six hundred and twenty three polynucleotides derived from a variety of human tissue sources.
PCT Publication No. WO 2003101283 relates to a combination which comprises many cDNAs and proteins
Japan Patent No. JP 2003259877 relates to many hepatic fibrosis disease markers.
Tzipora Shoshani, et al. Identification of a Novel Hypoxia-Inducible Factor 1-Responsive Gene, RTP801, Involved in Apoptosis. MOLECULAR AND CELLULAR BIOLOGY, April 2002, p. 2283-2293; this paper, co-authored by the inventor of the present invention, details the discovery of the RTP801 gene (a then novel HIF-1-dependent gene).
Anat Brafman, et al. Inhibition of Oxygen-Induced Retinopathy in RTP801-Deficient Mice. Invest Ophthalmol Vis Sci. 2004 October; 45 (10): 3796-805; also co-authored by the inventor of the present invention, this paper demonstrates that in RTP801 knock out mice, hypoxia does not cause degeneration of the retinal capillary network.
Leif W. Ellisen, et al. REDD1, a Developmentally Regulated Transcriptional Target of p63and p53, Links p63 to Regulation of Reactive Oxygen Species. Molecular Cell, Vol. 10, 995-1005, November, 2002; this paper demonstrates that overexpression of RTP801 (referred to therein as REDD1) leads to increased production of reactive oxygen species.
Richard D R, Berra E, and Pouyssegur J. Non-hypoxic pathway mediates the induction of hypoxia-inducible factor 1 alpha in vascular smooth muscle cells. J Biol. Chem. 2000, Sep. 1; 275(35): 26765-71. This paper demonstrates that HIF-1-dependent transcription may be induced by excessive production of reactive oxygen species.
Rangasami T, et al., Genetic ablation of Nrƒ2 enhances susceptibility to cigarette smoke-induced emphysema in mice. Submitted to Journal of Clinical Investigation. This work relates to mice with a compromised antoxidant defense (due to a germline inactivation of RTP801).
The mTOR Pathway
Tuberous sclerosis is an autosomal-dominant disorder caused by the mutation of one of the two tumor suppressor genes: TSC1 or TSC2, (TSC=Tuberous Sclerosis Complex) encoding protein products, hamartin, and tuberin, respectively. Both proteins form intracellular complexes exerting inhibitory activity on mammalian target of rapamycin (mTOR) kinase. It has been demonstrated that signal transduction from tuberin to mTOR is mediated by a G protein, Ras homologue enriched in brain (Rheb). In normal cells, tuberin having GTPase-activating protein properties toward Rheb controls signals of nutrient depletion, hypoxia, or stress, not allowing activation of mTOR and subsequent protein translation and cell proliferation. However, when environmental conditions change, tuberin is phosphorylated and it forms a complex with hamartin is degraded, and downstream targets of mTOR, S6K, and eEF2K, can be activated. (Jozwiak J, Jozwiak S, Grzela T, Lazarczyk M: Positive and negative regulation of TSC2 activity and its effects on downstream effectors of the mTOR pathway. Neuromolecular Med. 2005; 7(4):287-96.).
mTOR is a central regulator of protein synthesis the activity of which is modulated by a variety of signals. Energy depletion and hypoxia result in mTOR inhibition through a process involving the activation of AMP-activated protein kinase (AMPK) by LKB1 and subsequent phosphorylation of TSC2. It has been shown that mTOR inhibition by hypoxia requires the TSC1/TSC2 tumor suppressor complex and RTP801. Disruption of the TSC1/TSC2 complex through loss of TSC1 or TSC2 blocks the effects of hypoxia on mTOR, as measured by changes in the mTOR targets S6K and 4E-BP1, and results in abnormal accumulation of Hypoxia-inducible factor (HIF). In contrast to energy depletion, mTOR inhibition by hypoxia does not require AMPK or LKB1. Down-regulation of mTOR activity by hypoxia requires de novo mRNA synthesis and correlates with increased expression of RTP801. Disruption of RTP801 abrogates the hypoxia-induced inhibition of mTOR, and RTP801 overexpression is sufficient to down-regulate S6K phosphorylation in a TSC1/TSC2-dependent manner. (Brugarolas J, Lei K, Hurley R L, Manning B D, Reiling J H, Hafen E, Witters L A, Ellisen L W, Kaelin W G Jr.: Regulation of mTOR function in response to hypoxia by REDD1 and the TSC1/TSC2 tumor suppressor complex. Genes Dev. 2004 Dec. 1; 18(23):2893-904.)
Additionally, it has recently been demonstrated that RTP801 potently inhibit signaling through mTOR, working downstream of AKT and upstream of TSC2 to inhibit mTOR functions. (Corradetti M N, et al.,. J Biol Chem. 2005 Mar. 18; 280(11):9769-72.).